Amorphous calcium carbonate for treating a leukemia

ABSTRACT

The present invention provides compositions and methods of treating a leukemia, including chronic lymphocytic leukemia, wherein the method comprises administering a stabilized amorphous calcium carbonate to a person in need thereof.

FIELD OF THE INVENTION

The present invention relates to compositions and methods of treatingleukemia, and in particular chronic lymphocytic leukemia, usingamorphous calcium carbonate stabilized by at least one stabilizingagent.

BACKGROUND OF THE INVENTION

Chronic lymphocytic leukemia (CLL) is one of the most common types ofleukemia in adults. It often occurs during or after middle age and itrarely occurs in children. Usually, CLL does not cause any signs orsymptoms and is found during a routine blood test. CLL is commonlystaged according to the following scheme: stage 0−increased lymphocytescount (lymphocytosis) and appearance of “basket” cells without any othersigns (“basket” or “smudge” cells are essentially neoplastic cells thatgot “smudged” during slide preparation for routine blood smear becauseof the fragile nature of these cells); stage I−lymphocytosis andappearance of enlarged lymph nodes; stage II−lymphocytes+appearance ofenlarged liver or spleen and lymph nodes may be enlarged; stageIII−lymphocytes+decrease in red blood cells count, the liver, spleen orlymph nodes may be enlarged; and stage IV−lymphocytes+reduced plateletscount, lymph nodes, liver, or spleen may be enlarged and the number ofred blood cells may be reduced.

Whereas several methods for treating CLL are currently present atadvanced stages of the disease, at the early stages of the disease suchas stage 0, the treatment is usually a watchful waiting, i.e. closelymonitoring patient's condition without providing any treatment until thedisease reaches more advanced stage and/or until signs or symptomsappear or change.

Considering the constantly increasing incidence rate of CLL and improveddiagnostic tools allowing early diagnosis of the disease, there is anunmet need for new treatments, especially at the early stages of theCLL.

SUMMARY OF THE INVENTION

The present invention relates to methods of treating a leukemia, and inparticular chronic lymphocytic leukemia, using compositions comprisingstable amorphous carbonate. It has been unexpectedly found thatstabilized amorphous calcium carbonate may slow down or stop thedevelopment of chronic lymphocytic leukemia (CLL) at its initial stage.This is based on the observation that upon the administration of ACC bya person diagnosed as a zero stage CLL, the level of WBC (leucocytes)ceased elevating for at least three months following the beginning oftreatment.

In one aspect, the present invention provides a composition comprisingan amorphous calcium carbonate (ACC) for use in treatment of leukemia,wherein the ACC is stabilized by at least one stabilizing agent. In someembodiments, the leukemia is chronic leukemia and acute leukemia. Inother embodiments, the chronic leukemia is selected from chroniclymphocytic leukemia, chronic myelogenous leukemia and Hairy cellleukemia. According to one embodiment, the leukemia is chroniclymphocytic leukemia (CLL). According to certain embodiments, the CLL isat the early stage of the disease. According to some embodiment, theearly stage of CLL is a watchful waiting stage. According to anotherembodiment, the the early stage is stage 0.

In some embodiments, the leukemia is an acute leukemia selected fromacute myeloid leukemia and acute lymphoblastic leukemia.

According to certain embodiments, the ACC is selected from a natural ACCand synthetic ACC.

According to one embodiment, the ACC is a synthetic ACC. According tosuch embodiments, the ACC is stabilized by at least one stabilizer.According to some embodiments, the stabilizer is selected frompolyphosphate, organic acid, phosphorylated amino acid, bisphosphonate,phosphorylated organic acid, phosphoric or sulfuric esters of hydroxycarboxylic acid, hydroxyl bearing organic compound combined with alkalihydroxides, and any combination thereof. According to other embodiment,the polyphosphate is an inorganic polyphosphate selected fromtriphosphate, pyrophosphate and hexametaphosphate; the phosphorylatedamino acid is selected from phosphoserine or phosphothreonine; theorganic acid is selected from citric acid and tartaric acid; and thebisphosphonate is selected from alendronate, etidronic acid, zoledronicacid and medronic acid.

According to some embodiments, the treatment of a leukemia comprises atleast one of the following: ceasing, retarding, preventing and reversingthe development of the leukemia. According to some embodiments, ceasing,retarding or preventing the development of the leukemia comprisesceasing or retarding the progression of lymphocytosis.

According to other embodiments the treatment of a leukemia comprisesadministering about 500 mg/day to about 5000 mg/day of ACC. According tosome embodiments, the composition is orally administered.

According to one embodiment, the present invention provides acomposition comprising an amorphous calcium carbonate stabilized by atleast one stabilizer, for use in treating a chronic lymphocyticleukemia. According to some embodiments, the chronic lymphocyticleukemia is at stage 0 or at stage I of the disease. According to oneembodiment, the stabilizer is selected from phosphoserine, triphosphate,citric acid, a combination of phosphoserine and citric acid and acombination of triphosphate and citric acid.

According to some embodiments of the invention, the composition isformulated as a pharmaceutical composition, a nutraceutical composition,a food supplement or a medical food.

According to some embodiments, the present invention provides apharmaceutical composition comprising an amorphous calcium carbonate,for use in treating a chronic lymphocytic leukemia at stage 0, whereinthe ACC is stabilized by a stabilizer selected from phosphoserine,triphosphate, citric acid, a combination of phosphoserine and citricacid, and a combination of triphosphate and citric acid.

According to another aspect, the present invention provides a method oftreating a leukemia in a subject in need thereof comprisingadministering a composition comprising an effective amount of amorphouscalcium carbonate (ACC) stabilized by at least one stabilizing agent.According to some embodiments, the leukemia is a chronic leukemiaselected from the group consisting of chronic lymphocytic leukemia,chronic myelogenous leukemia, and Hairy cell leukemia, or an acuteleukemia selected from the group consisting of acute myeloid leukemiaand acute lymphoblastic leukemia. According to some embodiments, thepresent invention provides a method of treating chronic lymphocyticleukemia. According to one embodiment, the chronic leukemia is at stage0 of the disease.

According to some embodiments, the method of treating leukemia comprisesat least one of: ceasing, retarding or preventing the development of theleukemia. According to some embodiments ceasing, retarding or preventingthe development of the leukemia comprises ceasing or retarding theprogression of lymphocytosis. According to some embodiments, ceasing theprogression of lymphocytosis comprises maintaining the level of whiteblood cells at the same level for at least 3 months.

According to other embodiment, the method comprises administering about500 mg/day to about 5000 mg/day of ACC.

According to some embodiments, the composition comprising ACC and atleast stabilizer is formulated as a pharmaceutical composition ornutraceutical composition, a food supplement or a medical food.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to compositions and methods for treatmentof a leukemia. It has been surprisingly found, according to the presentinvention, that stable amorphous calcium carbonate (ACC) maysuccessfully slow down or even prevent further development of thechronic lymphocytic leukemia. Even more unexpected observation was thatthe progression of the disease had been inhibited at the very earlystage of the disease, at which the common acceptable treatment iswatchful waiting.

According to one aspect, the present invention provides a compositioncomprising an amorphous calcium carbonate (ACC) for use in treatment ofa leukemia, wherein the ACC is stabilized by at least one stabilizingagent.

The terms “treating” and “treatment” are uses herein interchangeably andrefer to taking steps to obtain beneficial or desired results, includingclinical results. Beneficial or desired clinical results include, butare not limited to, alleviation or amelioration of one or more symptomsassociated with leukemia and in particular chronic lymphocytic leukemia,delay or slowing the development or the progression of leukemia,amelioration, palliation or stabilization of the disease, and otherbeneficial results. In particular, according to one embodiment, treatinga leukemia comprises at least one of the following: ceasing, retardingreversing and preventing the development of the leukemia.

The term “leukemia” refers to a cancer of white blood cells (WBC)involving bone marrow and circulating WBCs, and of organs such as thespleen and lymph nodes. According to one embodiment, the leukemia is achronic leukemia. According to some embodiments, the chronic leukemia isselected from chronic lymphocytic leukemia, chronic myelogenousleukemia, and Hairy cell leukemia. According to other embodiments, theleukemia is an acute leukemia. According to some embodiments, the acuteleukemia is selected from acute myeloid leukemia and acute lymphoblasticleukemia.

According to some embodiments, the leukemia is a chronic leukemia. Thus,according to one embodiment, the present invention provides acomposition comprising an ACC stabilized by at least one stabilizingagent for use in treatment of chronic leukemia According to oneembodiment, the leukemia is a chronic lymphocytic leukemia (CLL). CLL iscommonly staged according to the following scheme: stage 0−increasedlymphocytes count (lymphocytosis) and appearance of “basket” cellswithout any other signs; stage I−lymphocytosis and appearance ofenlarged lymph nodes; stage II−lymphocytes+appearance of enlarged liveror spleen and lymph nodes may be enlarged, stageIII−lymphocytes+decrease in red blood cells count, the liver, spleen orlymph nodes may be enlarged; and stage IV−lymphocytes+reduced plateletscount, lymph nodes, liver, or spleen may be enlarged and the number ofred blood cells may be reduced. In one embodiment, CLL is at stage 0,stage I, stage II, stage III or at stage IV of the disease.

According to another embodiment, the CLL is at the early stage of thedisease. According to one embodiment, the early stage is a watchfulwaiting stage, i.e. the stage at which watchful waiting treatment iscommonly recommended. According to another embodiment, the early stageis stage 0 of CLL. According to another embodiment, the early stage isstage I of CLL.

The terms “amorphous calcium carbonate”, “ACC”, “stable ACC” and“stabilized ACC” are used herein interchangeably and refer to theamorphous form of calcium carbonate. The term “stable” as used hereinindicates that the calcium carbonate is maintained in the amorphous formfor a long period of time, for example for about at least 7 days in thesolid form having less than or about 30% crystalline calcium carbonate.According to any one of the above embodiments, the composition is stablefor at least 7 days. According to some embodiments, the composition isstable for at least 1 month. According to other embodiments, thecomposition is stable for at least 3 months. According to a furtherembodiment, the composition is stable for 6 months. According to certainembodiments, the composition is stable for at least 1 year. According toa particular embodiment, the composition is stable to at least 2 years.

According to any one of the above embodiments, the ACC stabilized by atleast one stabilizer is a natural ACC. The term “natural ACC” as usedherein refers to any ACC isolated or derived from a natural source.Non-limiting examples of natural sources of ACC include gastroliths offreshwater crustaceans. In certain embodiments, the naturally occurringACC source includes gastrolith organs, or a portion thereof ground to afine powder, essentially as described in WO 2005/115414. Optionally, ACCcomprises a combination of naturally occurring and synthesized ACC.

According to another embodiment, the ACC is a synthetic ACC. The term“synthetic ACC” as used herein generally refers to any ACC produced byman ex-vivo. According to some embodiments, the ACC is a synthetic ACCstabilized by at least one stabilizer as defined herein below.

The terms “stabilizing agent” and “stabilizer” are used hereininterchangeably and refer to any substance that contributes topreserving calcium carbonate in the amorphous state during ACCproduction, formulating and/or storage. In certain embodiments, thestabilizing agent is a single agent. In other embodiments, use ofseveral stabilizing agents is encompassed.

ACC Stabilizers

The stabilizer may comprise a molecule having one or more functionalgroups selected from, but not limited to, hydroxyl, carboxyl, ester,amine, phosphino, phosphono, phosphate, sulfonyl, sulfate or sulfinogroups. The hydroxy bearing compounds, combined with the hydroxide,optionally also bear other functions like carboxyl, etc. but with thehydroxyl not being esterified.

According to some embodiments, the stabilizer has low toxicity or notoxicity to mammalian cells or organism, and in particular to a humanbeing. According to some embodiments, the stabilizer is of food,nutraceutical or pharmaceutical grade.

In certain embodiments, the ACC stabilizing agent is independently ateach occurrence, an organic acid, phosphorylated, phosphonated, sulfatedor sulfonated organic compound, phosphoric or sulfuric ester of ahydroxyl carboxylic acid, an organoamine compound, an organic compoundcomprising a hydroxyl, an organophosphorous compound or a salt thereof,phosphorylated amino acids and derivatives thereof, a bisphosphonatecompound, an organophosphate compound, an organophosphonate compound, aninorganic phosphorous acid, an organic compound having multiplefunctional groups as defined above, an inorganic phosphate andpolyphosphate compound, an organic compound having a polyphosphatechain, an organic surfactant, a bio-essential inorganic ion, or anycombination thereof.

According to some embodiments, the stabilizer is an organic acid.According to certain embodiments, the organic acid is selected fromascorbic acid, citric acid, lactic acid, acetic acid, oxalic acid,malonic acid, glutaconic acid, succinic acid, maleic acid, lactic acid,aconitic acid, and optionally include compounds having at least twocarboxylic groups and molecular weight not larger than 250 g/mol, suchas citric acid, tartaric acid, malic acid, etc. According to oneparticular embodiment, the stabilizer is citric acid.

In another embodiment, the phosphoric ester of hydroxyl carboxylic acidsis a phosphoenolpyruvate. In another embodiment, the phosphoric orsulfuric esters of hydroxyl carboxylic acids comprise amino acids, e.g.phosphorylated amino acids. Examples of such esters are phosphoserine,phosphothreonine, sulfoserine, sulfothreonine and phosphocreatine.

The hydroxyl bearing compounds combined with hydroxide may comprise, forexample, mono-, di- tri-, oligo-, and polysaccharides like sucrose orother polyols like glycerol. The hydroxyl bearing compounds may furthercomprise hydroxy acids like citric acid, tartaric acid, malic acid,etc., or hydroxyl-bearing amino acids such as serine or threonine. Eachpossibility represents a separate embodiment, of the present invention.

Some specific unlimited examples for such ACC stabilizers include phyticacid, citric acid, sodium pyrophosphate dibasic, adenosine5′-monophosphate (AMP) sodium salt, adenosine 5′-diphosphate (ADP)sodium salt and adenosine 5′-triphosphate (ATP) disodium salt hydrate,phosphoserine, phosphorylated amino acids, food grade surfactants,sodium stearoyl lactylate, and combinations thereof.

According to some embodiments, the stabilizer comprises at least onecomponent selected from phosphoric or sulfuric esters of hydroxylcarboxylic acids, such as phosphoenolpyruvate, phosphoserine,phosphorthreonine, sulfoserine or sulfothreonine and hydroxyl bearingorganic compounds, selected from mono-, di-, tri-, oligo- andpolysaccharides, for example, sucrose, mannose, glucose. The hydroxylbearing compound may further comprise at least one alkali hydroxide,such as sodium hydroxide, potassium hydroxide and the like. Thephosphorylated acids may be present in oligopeptides and polypeptides.In other embodiments, of the invention, the stabilizer is an organicacid selected from monocarboxylic acid or multiple carboxylic acid, e.g.dicarboxylic acid or tricarboxylic acid. Each possibility represents aseparate embodiment, of the invention. The organic acid may be asdefined above.

In some embodiments, of the invention, the ACC stabilizer is selectedfrom phosphorylated amino acids, polyols and combinations thereof. Insome embodiments, the stable ACC comprises a phosphorylated compound asa stabilizer wherein the phosphorylation is performed on the hydroxylgroup of an organic compound. In some embodiments, the stable ACCcomprises a stabilizer selected from the group consisting of citricacid, phosphoserine, phosphothreonine and combinations thereof. Thenon-limiting examples of stabilizers containing phosphate, phosphite,phosphonate groups and salts or esters thereof include phytic acid,dimethyl phosphate, trimethyl phosphate, sodium pyrophosphate,tetraethyl pyrophosphate, ribulose bisphosphate, etidronic acid andother medical bisphosphonates, 3-phosphoglyceric acid salt,glyceraldehyde 3-phosphate, 1-deoxy-D-xylulose-5-phosphate sodium salt,diethylene triamine pentakis(methylphosphonic acid),nitrilotri(methylphosphonic acid), 5-phospho-D-ribose 1-diphosphatepentasodium salt, adenosine 5′-diphosphate sodium salt, adenosine5′-triphosphate disodium salt hydrate, α-D-galactosamine 1-phosphate,2-phospho-L-ascorbic acid trisodium salt, α-D-galactose 1-phosphatedipotassium salt pentahydrate, α-D-galactosamine 1-phosphate,O-phosphorylethanolamine, disodium salt hydrate,2,3-diphospho-D-glyceric acid pentasodium salt, phospho(enol)pyruvicacid monosodium salt hydrate, D-glyceraldehyde 3-phosphate, sn-glycerol3-phosphate lithium salt, D-(−)-3-phosphoglyceric acid disodium salt,D-glucose 6-phosphate sodium salt, phosphatidic acid, ibandronate sodiumsalt, phosphonoacetic acid, DL-2-amino-3-phosphonopropionic acid orcombinations thereof. The bio-essential inorganic ions may include,inter alia, Na, K, Mg, Zn, Fe, P, S, N, P or S in the phase of oxides,or N as ammonia or nitro groups.

According to some embodiments, the stabilizer is a polyphosphate orpharmaceutically acceptable salts thereof. According to someembodiments, the polyphosphate is physiologically compatible, watersoluble polyphosphate salt selected from the group consisting of sodium,potassium and any other essential cation of polyphosphate. In oneembodiment, the polyphosphate is organic or inorganic polyphosphate. Theterm “polyphosphate” as used herein refers to polymeric esters of PO4.According to some embodiments, the polyphosphate is physiologicallycompatible water soluble polyphosphate salt selected from the groupconsisting of sodium and potassium polyphosphate. In some embodiments,the polyphosphate is an inorganic polyphosphate or pharmaceuticallyacceptable salts thereof. Not-limiting examples of such salt are Na, K,Mg, Mn and Zn. According to some embodiments, the inorganic phosphatecomprise 2 to 10 phosphate groups, e.g. 2, 3, 4, 5, 6, 7, 8, 9, or 10phosphate group. According to some embodiments, the polyphosphate isselected from pyrophosphate, triphosphate, and hexametaphosphate.According to one embodiment, the stabilizer is pyrophosphate orpharmaceutically acceptable salts thereof such as sodium pyrophosphate.According to another embodiment, the stabilizer is triphosphate orpharmaceutically acceptable salts thereof such as sodium triphosphate.The term “triphosphate” and “tripolyphosphate” are used hereininterchangeably. According to a further embodiment, the stabilizer ishexametaphosphate or a pharmaceutically acceptable salt thereof suchsodium hexametaphosphate.

According to some embodiments, the stabilizer is a bisphosphonate orpharmaceutically acceptable salts thereof. The not-limiting examples ofsalt are Na, K, Mg, Mn and Zn.

The term “bisphosphonate” as used herein refers to organic compoundshaving two phosphonate (PO(OH)2) groups. The term further relates tocompounds having a backbone of PO3-organic-PO3. Most typical is a seriesof bisphosphonates that are used as pharmaceuticals for treatingosteoporosis. According to some embodiments, the bisphosphonate isselected from the group consisting of etidronic acid, zoledronic acid,medronic acid, alendronic acid and a pharmaceutically acceptable saltthereof. According to some embodiments, the stabilizer is an etidronicacid or a pharmaceutically acceptable salt thereof. According to anotherembodiment, the stabilizer is a zoledronic acid or a pharmaceuticallyacceptable salt thereof. According to a further embodiment, thestabilizer is a medronic acid or a pharmaceutically acceptable saltthereof. According to certain embodiments, the stabilizer is alendronicacid or a pharmaceutically acceptable salt thereof.

According to certain embodiments, the stabilizer is a phosphorylatedamino acid. According to one embodiment, the phosphorylated amino acidis phosphoserine. According to another embodiment, the phosphorylatedamino acid is phosphothreonine.

According to some embodiments, the stabilizer is polyphosphate or abisphosphonate as defined hereinabove, and the molar ratio between Patoms of the stabilizer and Ca atoms of the ACC (P:Ca molar ratio) isabout 1:90 to 1:1. In one embodiment, the P:Ca molar ratio is about 1:40to about 1:1. In a further embodiment, the P:Ca molar ratio is about1:35 to about 1:2. In certain embodiments, the P:Ca molar ratio is about1:30 to about 1:3. In certain embodiments, the P:Ca molar ratio is about1:28 to about 1:3. In other embodiment, the P:Ca molar ratio is about1:25 to about 1:4. In further embodiment, the P:Ca molar ratio is about1:20 to about 1:5. In another embodiment, the P:Ca molar ratio is about1:20 to about 1:6. In a particular embodiment, the P:Ca molar ratio isabout 1:15 to about 1:5. In another particular embodiment, the P:Camolar ratio is about 1:25 to about 1:5. According to some embodiments,such polyphosphate is pyrophosphate, triphosphate, hexametaphosphate ora pharmaceutically acceptable salt thereof. According to anotherembodiment, the bisphosphonate is alendronic acid, etidronic acid,zoledronic acid or medronic acid and the P:Ca molar ratio is as definedhereinabove.

According to some embodiments, the calcium content (Ca content) of suchcompositions comprising polyphosphate or bisphosphonate as a stabilizeris about 1 wt % to about 39 wt %, about 5 wt % to about 39 wt %, about10% to about 39 wt %, about 15% to about 39 wt %, about 20 wt % to about38 wt %, about 25 wt % to about 38 wt %, or about 30 to about 38. Theterms “Ca content” and “calcium content” is used herein interchangeablyand refer to the content of calcium of the ACC in the final composition.

In certain embodiments, the P:Ca molar ratio is about 1:40 to about 1:1,and the Ca content is about 20 wt % to about 39 wt %. In someembodiments, the molar ratio is 1:28 to about 1:3, and the Ca content isabout 30 wt % to about 38 wt %. In another embodiment, the molar ratiois 1:25 to about 1:5, and the Ca content is about 30 wt % to about 36 wt%.

According to some embodiments, the stabilizer is selected from the groupconsisting of a polyphosphate, phosphorylated amino acid,bisphosphonate, citric acid, tartaric acid and any combination thereof.According to one embodiment, the polyphosphate is selected from thegroup consisting of triphosphate, pyrophosphate, and hexametaphosphate,the phosphorylated amino acid is phosphoserine or phosphothreonine, andthe bisphosphonate is selected from the group consisting of alendronate,etidronic acid, zoledronic acid and medronic acid.

According to some embodiments, the stabilizer is polyphosphate orbisphosphonate and the molar ratio between P atoms of the stabilizer andCa atoms of the ACC is about 1:90 to 1:1.

The stabilized ACC may be stabilized by more than one stabilizers, e.g.two or more stabilizers. In some embodiments, more than one stabilizers,e.g. 2, 3 or 4 stabilizers are added. In some embodiments, the firststabilizer and the second stabilizer are similar. In other embodiments,the first stabilizer and the second stabilizer are differentstabilizers. The first and the second stabilizers may be eachindependently as defined hereinabove. The stable ACC can comprise morethan two stabilizers, wherein one or more stabilizers are added to theACC during the formation and precipitation of the ACC.

According to one embodiment, ACC is stabilized by a combination ofphosphoserine and citric acid. According to another embodiment, ACC isstabilized by a combination of triphosphate and citric acid.

According to any one of the above embodiments, the stabilized ACC ispresent is the form of a powder. According to some embodiments, theparticles of the ACC in the powder have a particle size of less thanabout 100 μm. In some embodiments, the ACC particles have a particlesize of about 100 μm to about 5 μm. In other embodiments, the particlesize is about 50 μm to about 5 μm, or about 30 to about 5 μm. In oneparticular embodiments the particles having the size less than 50 μm,less than 40 μm, less than 30 μm, less than 20 μm or less than 10 μm.According to some embodiments at least 70%, at least 80% or at least 90%of the ACC particles have the particle size of less than 5 μm.

The term “particle” as used herein refers to a discrete microparticle ora nanoparticle of ACC stabilized by the stabilizer as definedhereinabove, as well as to the aggregates or agglomerates thereof.According to some embodiments, the particles are primary particles ofthe stabilized ACC. The basic nanoparticles are in the range of 5 to 500nm or 10 to 300 nm or 20 to 100 nm. These nanoparticles immediatelyagglomerate and aggregate into much larger secondary particles. Theseaggregation and agglomeration can be then broken by milling anddissolution techniques into smaller particles. According to otherembodiments the particles are agglomerates or aggregates of the primaryparticles, i.e. secondary particles.

According to any one of the above embodiments, the term treating aleukemia comprises at least one of the following: ceasing, retardingpreventing or reversing the development of the leukemia. According toone embodiment, treating a leukemia comprises ceasing the development ofthe leukemia. The development of the disease is characterized by changein at least one of the parameters associated with or characterizingleukemia. According to some embodiments, treating a leukemia comprisesretarding the development of the leukemia. According to a furtherembodiment, treating a leukemia comprises preventing the development ofthe leukemia. According to one embodiment, the leukemia is CLL. Thus, inone embodiment, treating CLL comprises ceasing, retarding or preventingthe development of CLL. The development of the disease is characterizedby a change in at least one of parameters associated or characterizingCLL. Non-limiting examples for such parameters are increase in WBC count(lymphocytosis), appearance of “basket” cells, appearance of enlargedlymph nodes liver and/or spleen, reduction in red blood cells (RBC)count, and reduction in platelets count. Thus, ceasing, retarding orslowing down a change in any one of these parameters is considered to orindicates for ceasing, retarding, slow down or preventing thedevelopment of CLL. According to one embodiment, treating of CLLcomprises ceasing, retarding or preventing development of CLL at earlystages of the disease. According to a further embodiment, the earlystage is stage 0. According to another embodiment, the early stage isstage I. In certain embodiments, treating of CLL comprises treating atstage 0 and/or at stage I. According to one embodiment, treating CLL,e.g. ceasing, retarding preventing or reversing the development of theCLL comprises ceasing, retarding or reversing the progression oflymphocytosis. According to some embodiments, retarding or reversing theprogression of lymphocytosis comprises maintaining the level oflymphocyte at the same level for at least 2 months. According to anotherembodiment, retarding or reversing the progression of lymphocytosiscomprises maintaining the level of lymphocyte at the same level for atleast 3, 4, 5, 6, 9 or 12 months. According to another embodiment,treating CLL comprises preventing the appearance, ceasing or retardingthe progression of appearance of “basket” cells, or reduction in thenumber of “basket” cells. According to another embodiment, treating theCLL comprises preventing the appearance of enlarged lymph nodes liverand/or spleen; or ceasing or retarding the enlargement of lymph nodesliver and/or spleen. According to one embodiment, treating the CLLcomprises obtaining a decrease in the size of the enlarged lymph nodesliver and/or spleen. According to one embodiment, the decrease in thesize is of 10%, 20%, 30%, 40% or 50%. According to another embodiment,treating CLL comprises preventing, ceasing or retarding the decrease inRBC count, platelets count or both. According to some embodiments,ceasing or retarding the decrease in RBC count, platelets count or bothcomprises maintaining the level of RBC, platelets or both at the samelevel for at least 2, 3, 4, 5, 6, 9 or 12 months. According to a furtherembodiment, treating CLL comprises increase in RBC count and/or inplatelets count. Monitoring the development of the CLL and in particularthe parameters characterizing CLL, may be obtained by any known method.

The composition for use of the present invention may be administered byany known method. The terms “administering” or “administration of” thecomposition of the present invention can be carried out using one of avariety of methods known to those skilled in the art. For example, thecomposition may be administered enterally or parenterally. Enterallyrefers to administration via the gastrointestinal tract including peros, sublingually or rectally. Parenteral administration includesadministration intravenously, intradermally, intramuscularly,intraperitoneally, subcutaneously, ocularly, sublingually, intranasally,by inhalation, intraspinally, intracerebrally, and transdermally (byabsorption, e.g., through a skin duct). The composition of the presentinvention can also appropriately be introduced by rechargeable orbiodegradable polymeric devices or other devices, e.g., patches andpumps, or formulations, which provide for the extended, slow orcontrolled release of the compound or agent. Administering can also beperformed, for example, once, a plurality of times, and/or over one ormore extended periods. In some aspects, the administration includes bothdirect administration, including self-administration, and indirectadministration, including the act of prescribing a drug or a medicalfood. For example, as used herein, a physician who instructs a patientto self-administer a drug or a medical food, or to have the drug or themedical food administered by another and/or who provides a patient witha prescription for a drug or a medical food is administering the drug ora medical food to the patient.

In one embodiment, the composition of the present invention isadministered via a systemic administration. For example, the compositionof the present invention may be administered orally, sublingually orrectally. Alternatively the composition of the present invention may beadministered intravenously, intradermally, intramuscularly,intraperitoneally, subcutaneously, ocularly, sublingually, intranasally,by inhalation, intraspinally, intracerebrally, and transdermally. In onespecific embodiment, the composition of the present invention isadministered orally.

According to any one of the above embodiments, treating leukemiacomprises administering about 500 mg/day to about 5000 mg/day, about 750mg/day to about 4000 mg/day of ACC, about 1000 mg/day to about 3000mg/day, or about 1500 mg/day to about 2500 mg/day of stabilized ACC.According to one embodiment, treating leukemia comprises administeringabout 500 mg/day to about 3000 mg/day or about 1000 mg/day to about 2500mg/day of stabilized ACC. According to one embodiment, the leukemia isCLL.

According to any one of the above embodiments, the composition of thepresent invention is formulated as a pharmaceutical composition,nutraceutical composition, a food supplement or a medical food.

According to one embodiment, the composition comprising a stabilized ACCfor use in treatment of a leukemia is a pharmaceutical composition.According to another embodiment, the composition comprising a stabilizedACC for use in treatment of a leukemia is a nutraceutical composition.According to yet another embodiment, the composition comprising astabilized ACC for use in treatment of a leukemia is a food supplement.According to a further embodiment, the composition comprising astabilized ACC for use in treatment of a leukemia is a medical food.According to such embodiments, the leukemia is CLL. According to anotherembodiment, the CLL is at stage 0 or stage I of the disease.

The term “pharmaceutical composition” as used herein refers to anycomposition comprising a stabilized ACC and a pharmaceuticallyacceptable excipient.

The term “pharmaceutically acceptable carrier” or “pharmaceuticallyacceptable excipient” as used herein refers to any and all solvents,dispersion media, preservatives, antioxidants, coatings, isotonic andabsorption delaying agents, surfactants, fillers, disintegrants,binders, diluents, lubricants, glidants, pH adjusting agents, bufferingagents, enhancers, wetting agents, solubilizing agents, surfactants,antioxidants the like, that are compatible with pharmaceuticaladministration. The use of such media and agents for pharmaceuticallyactive substances is well known in the art. The compositions may containother active compounds providing supplemental, additional, or enhancedtherapeutic functions.

The terms “pharmaceutically acceptable” and “pharmacologicallyacceptable” include molecular entities and compositions that do notproduce an adverse, allergic, or other untoward reactions whenadministered to an animal, or human, as appropriate. For humanadministration, preparations should meet sterility, pyrogenicity,general safety, and purity standards as required by a government drugregulatory agency, e.g., the United States Food and Drug Administration(FDA) Office of Biologics standards.

As used herein, the term “nutraceutical composition” refers to acomposition suitable for use in human beings or animals, comprising oneor more natural products with therapeutic action which provide a healthbenefit or have been associated with disease prevention or reduction.

The term “food supplement” is used to mean a product containing saidcomposition and intended to supplement the food by providing nutrientsthat are beneficial to health according to any acceptable directive,such as European directive. For example, a food supplement may be acapsule or a tablet for swallowing, or a powder or small vial to mixwith a food and providing beneficial health effects.

As used herein, the term “medical food” refers to a food item speciallyformulated for the dietary management of a disease or disorder in asubject.

The composition of the present invention may be formulated as definedhereinabove. In one particular embodiment, the composition comprising astabilized ACC is formulated as a solid dosage form selected fromtablets, capsules, powder or granules. In another embodiment, thecomposition comprising a stabilized ACC is formulated as a liquid orsemi-liquid dosage form selected from an elixir, tincture, suspension,syrup, emulsion or gel. According to a further embodiment, thecomposition is formulated as a composition for inhalation.

According to certain embodiments, the present invention provides acomposition comprising an amorphous calcium carbonate stabilized by atleast one stabilizer, for use in treating a leukemia. According to oneembodiment, the leukemia is a chronic leukemia such as chroniclymphocytic leukemia, chronic myelogenous leukemia, and Hairy cellleukemia. According to one embodiment, the present invention provides acomposition comprising an ACC for use in treatment of CLL, wherein theACC is stabilized by at least one stabilizing agent. According to oneembodiment, the treating comprises treating CLL at stage 0 of thedisease. According to another embodiment, the treating comprisestreating at stage I of the disease. According to a further embodiment,treating comprises administering about 500 mg/day to about 5000 mg/day,or about 750 mg/day to about 4000 mg/day, about 1000 mg/day to about3000 mg/day, or about 1500 mg/day to about 2500 mg/day of stabilizedACC. According to some embodiments, the stabilizing agent is selectedfrom the group consisting of a polyphosphate such as triphosphate,pyrophosphate, and hexametaphosphate; phosphorylated amino acid such asphosphoserine or phosphothreonine, bisphosphonate such as alendronate,etidronic acid, zoledronic acid and medronic acid; organic acid such ascitric acid and tartaric acid; and any combination thereof. According toone embodiment, treating a leukemia comprises at least one of ceasing,retarding and preventing the development of the leukemia. According toone embodiment, the composition is a pharmaceutical composition.According to another embodiment, the composition is nutraceuticalcomposition or food supplement.

According to one embodiment, the present invention provides apharmaceutical composition comprising an ACC and a pharmaceuticallyacceptable excipient, for use in treatment of CLL at stage 0 and/orstage I of the disease, wherein the ACC is stabilized by a stabilizerselected from phosphoserine; triphosphate; citric acid; a combination ofphosphoserine and citric acid; and a combination of triphosphate andcitric acid. According to one embodiment, the treatment comprisesadministration of about 500 mg/day to about 5000 mg/day or about 1000mg/day to about 3000 mg/day of stabilized ACC. According to oneembodiment, the pharmaceutical composition is orally administered.

According to one embodiment, the present invention provides a foodsupplement comprising an ACC, for use in treatment of CLL at stage 0and/or stage I of the disease, wherein the ACC is stabilized by astabilizer selected from phosphoserine, triphosphate, citric acid, acombination of phosphoserine and citric acid; and a combination oftriphosphate and citric acid. According to one embodiment, the treatmentcomprises administration of about 500 mg/day to about 5000 mg/day orabout 1000 mg/day to about 3000 mg/day of stabilized ACC.

According to one aspect the present invention provides a method oftreating a leukemia in a subject in need thereof comprisingadministering a composition comprising an effective amount of amorphouscalcium carbonate (ACC) stabilized by at least one stabilizing agent.According to one embodiment, the leukemia is a chronic leukemia.According to some embodiments, the chronic leukemia is selected fromchronic lymphocytic leukemia, chronic myelogenous leukemia, and Hairycell leukemia. According to other embodiments, the leukemia is an acuteleukemia. According to some embodiments, the acute leukemia is selectedfrom acute myeloid leukemia and acute lymphoblastic leukemia.

The term “effective amount” as used herein refers to a sufficient amountof the composition comprising stabilized ACC for treating leukemia at areasonable benefit/risk ratio applicable to any medical or nutritionaltreatment.

According to one embodiment, the present invention provides a method oftreating CLL in a subject in need thereof comprising administering acomposition comprising an effective amount of amorphous calciumcarbonate (ACC) stabilized by at least one stabilizing agent as definedabove. According to one embodiment, the treating is at the early stagesof the disease e.g. at stage 0. According to a further embodiment, themethod comprises administering about 500 mg/day to about 5000 mg/day,about 750 mg/day to about 4000 mg/day, about 1000 mg/day to about 3000mg/day, or about 1500 mg/day to about 2500 mg/day of stabilized ACC.According to one embodiment, treating CLL comprises at least one of thegroups consisting of ceasing, retarding and preventing the developmentof the leukemia. According to some embodiments, treating a CLL compriseas least one of the group ceasing, retarding or reversing theprogression of lymphocytosis; ceasing or retarding the progression ofappearance of “basket” cells; reduction in the number of “basket” cells;preventing the appearance of enlarged lymph nodes liver and/or spleen;ceasing or retarding the enlargement of lymph nodes liver and/or spleen;decreasing the size of the enlarged lymph nodes liver and/or spleen;preventing, ceasing or retarding the decrease in RBC count; preventing,ceasing or retarding the decrease in platelets count; increase in RBCcount; and increase in RBC in platelets count. According to oneembodiment, retarding or reversing the progression of lymphocytosiscomprises maintaining the level of lymphocyte at the same level for atleast 3, 4, 5, 6, 9 or 12 months. According to another embodiment, thedecrease in the size of the enlarged lymph nodes liver and/or spleencomprises a decrease of 10%, 20%, 30%, 40% or 50%. According to otherembodiments, ceasing or retarding the decrease in RBC count, plateletscount or both comprises maintaining the level of RBC, platelets or bothat the same level for at least 2, 3, 4, 5, 6, 9 or 12 months. Accordingto some embodiments, the stabilizing agent is selected from the groupconsisting of a polyphosphate such as triphosphate, pyrophosphate, andhexametaphosphate; phosphorylated amino acid such as phosphoserine orphosphothreonine, bisphosphonate such as alendronate, etidronic acid,zoledronic acid and medronic acid; organic acid such as citric acid andtartaric acid; and any combination thereof. According to any one of theabove embodiments, the composition of the present invention isformulated as a pharmaceutical or nutraceutical composition, a foodsupplement or a medical food. In one embodiment, the stabilized ACC orthe composition is administered via a systemic administration. Forexample, the composition is administered orally, sublingually orrectally. Alternatively the composition is administered intravenously,intradermally, intramuscularly, intraperitoneally, subcutaneously,ocularly, sublingually, intranasally, by inhalation, intraspinally,intracerebrally, and transdermally. In one embodiment, the compositionis administered orally. According to one embodiment, the methodcomprises administering a pharmaceutical composition comprising stableACC as described above.

According to another aspect, the present invention provides amorphouscalcium carbonate stabilized by at least one stabilizer for preparing amedicament for use in treating a leukemia. According to one embodiment,the leukemia is a chronic leukemia such as chronic lymphocytic leukemia,chronic myelogenous leukemia, and Hairy cell leukemia or an acuteleukemia such as acute myeloid leukemia and acute lymphoblasticleukemia. According to one particular embodiment, the leukemia is CLL.According to one embodiment, the treating comprises treating at theearly stages of the disease e.g. at stage 0. According to a furtherembodiment, the method comprises administering about 500 mg/day to about5000 mg/day, about 750 mg/day to about 4000 mg/day of ACC, about 1000mg/day to about 3000 mg/day, or about 1500 mg/day to about 2500 mg/dayof stabilized ACC. According to some embodiments, the stabilizing agentis selected from the group consisting of a polyphosphate such astriphosphate, pyrophosphate, and hexametaphosphate; phosphorylated aminoacid such as phosphoserine or phosphothreonine, bisphosphonate such asalendronate, etidronic acid, zoledronic acid and medronic acid; organicacid such as citric acid and tartaric acid; and any combination thereof.

The terms “comprising”, “comprise(s)”, “include(s)”, “having”, “has”,and “contain(s)” are used herein interchangeably and have the meaning of“consisting at least in part of′. When interpreting each statement inthis specification that includes the term “comprising”, features otherthan that or those prefaced by the term may also be present. Relatedterms such as “comprise” and “comprises” are to be interpreted in thesame manner. The terms “have”, “has”, having” and “comprising” may alsoencompass the meaning of “consisting of” and “consisting essentiallyof”, and may be substituted by these terms. The term “consisting of”excludes any component, step or procedure not specifically delineated orlisted. The term “consisting essentially of” means that the compositionor component may include additional ingredients, but only if theadditional ingredients do not materially alter the basic and novelcharacteristics of the claimed compositions or methods.

As used herein, the term “about”, when referring to a measurable valuesuch as an amount, a temporal duration, and the like, is meant toencompass variations of +/−10%, or +/−5%, +/−1%, or even +/−0.1% fromthe specified value.

The term “or” as used herein, denotes alternatives that may, whereappropriate, be combined; that is, the term “or” includes each listedalternative separately as well as their combination if the combinationis not mutually exclusive.

Having now generally described the invention, the same will be morereadily understood through reference to the following examples, whichare provided by way of illustration and are not intended to be limitingof the present invention.

Examples

A 72 years old man, generally healthy, showed a graduate increase inleucocytes values in several consequent blood tests. In addition,“basket cells” were observed and the person showed a reduction in thevitality; the man tended to fall asleep frequently.

No other symptoms that may be attributed to chronic lymphocytic leukemia(CLL) were observed. The person was diagnosed as stage 0 CLL patient andstarted a treatment with amorphous calcium carbonate (ACC) at Nov. 20,2015 (five days after the blood test) according to the followingregiment:

During the first three weeks: 3 Density® tablets/day orally (Density® toAmorphical Ltd.).

After three weeks, the daily dose was elevated to 6 Density®tablets/day.

Each Density® tablet contains 666 mg ACC as API (i.e. AmorphousCaCO3+Aerosil+drug substance stabilizers) which corresponds to 500 mgCaCO3, and equivalent to 200 mg elemental calcium (hereinafter the doserefers to the amount of elemental calcium).

The results of several consecutive blood tests (before and after thebeginning of ACC administration) are presented in Tables 1 and 2.

TABLE 1 Full blood count values in several consecutive blood test test25 Mar. 2015 20 Aug. 2015 15 Nov. 2015 7 Dec. 2015* 17 Feb. 2016* Normalrange WBC 25.9 38.8 52.1 55.4 50.3  4.5-11 (leucocytes) RBC 4.73 4.915.24 4.59  4.5-5.5 hemoglobin 15 15.4 16.1 14.4 13.5-18 hematocrit 43.446.2 49.3 43.6   41-53% MCV 92 94 94 95   79-97 MCH 31.7 31.4 30.7 31.4  27-34 MCHC 34.6 33.3 32.7 33   32-36 RDW 13.4 14.1 14.2 14.1 11.6-15platelets 146 139 144 149 141  150-450 MPV- 12.5 13 12.4 11.9  8.5-12.9Neutrophils % 9.8 19 6 13 18   40-75 confirm Lymphocytes % 83.1 72 91 8477   22-44 confirm Monocytes % 8 2 3 4   3-13 confirm eosinophils 1 0 01   0-6 basophils 0 1 0 0   0-2 presence of X X X X X 0 basket cellsPH(u) — — — 6.5   5-8 nitrite — — — — ketone — — — — glucose — — — —urobilinogen — — — — protein — — — — leucocytes — — — — erythrocytes — —— 10   0-10 Immunoglobulin 32.7 34.7 38.3 30.5   40-230 mg/dl MImmunoglobulin 731.9 817.5 832.5 775.8  737-1607 mg/dl G Immunoglobulin199.8 210.7 209.8 199.9   80-520 mg/dl A RBC-red blood cells; MCV-meancell volume; MCHC-M.cell HB count; RDW-red cell distri.width; MPV-meanplatelet volume. *the were tests performed after the beginning of ACCadministration.

TABLE 2 The WBC count in several consecutive tests February June JulyMarch August 15 Nov. *December *February Normal 2013 2014 2014 2015 20152015 2015 2016 range WBC 13.8 25 22.7 25.9 38.8 52.1 55.4 50.3 4.5-11 %Lymphocytes 83.1 72.6 96 84 77  22-44 % Monocytes 3 4 8 2 3 4   3-13 %Neutrophils 9.8 19 6 13 18  40-75 *the tests were performed after thebeginning of ACC administration.

It can be clearly seen that the elevation in WBC count was stopped afterthe patient started administrating ACC, and actually, the WBC count wasreduced in the last blood test. In addition, an improvement in severalother parameters, e.g. an increase in myocytes and neutrophils count wasobserved. Moreover, the patient became much more vital; the person didno longer tend to fall asleep as frequently as before the treatment.

As follows from these results, administration of ACC allows preserving asubstantially constant level of WBC indicating that the treatmenteffectively prevents further progression of CLL and maintains it at zerolevel.

Although the present invention has been described herein above by way ofpreferred embodiments thereof, it can be modified, without departingfrom the spirit and nature of the subject invention as defined in theappended claims.

1. A method of treating a leukemia in a subject in need thereof, themethod comprising: administering a composition including an effectiveamount of amorphous calcium carbonate (ACC) stabilized by at least onestabilizing agent.
 2. The method of claim 1, wherein the leukemia is achronic leukemia selected from the group consisting of chroniclymphocytic leukemia, chronic myelogenous leukemia, and Hairy cellleukemia, or an acute leukemia selected from the group consisting ofacute myeloid leukemia and acute lymphoblastic leukemia.
 3. The methodof claim 2, wherein the leukemia is a chronic lymphocytic leukemia. 4.The method of claim 1, wherein the at least one stabilizing agent isselected from the group consisting of polyphosphate, organic acid,phosphorylated amino acids, bisphosphonate, phosphorylated organic acid,phosphoric or sulfuric ester of hydroxy carboxylic acid, hydroxylbearing organic compound combined with alkali hydroxides and anycombination thereof.
 5. The method of claim 4, wherein the polyphosphateis an inorganic polyphosphate selected from the group consisting oftriphosphate, pyrophosphate. and hexametaphosphate; the phosphorylatedamino acid is selected from the group consisting of phosphoserine orphosphothreonine; the organic acid is selected from the group consistingof citric acid and tartaric acid; and the bisphosphonate is selectedfrom the group consisting of alendronate, etidronic acid, zoledronicacid, and medronic acid.
 6. The method of claim 5, wherein the at leastone stabilizing agent is a polyphosphate or bisphosphonate, and whereinthe molar ratio between P atoms of the stabilizing agent and Ca atoms ofthe ACC is about 1:90 to 1:1.
 7. The method of claim 1, wherein treatinga leukemia includes at least one of ceasing, retarding, reversing, orpreventing the development of the leukemia.
 8. The method of claim 7,wherein ceasing, retarding, reversing, or preventing the development ofthe leukemia includes ceasing, retarding, or reversing the progressionof lymphocytosis.
 9. The method of claim 8, wherein ceasing theprogression of lymphocytosis includes preserving the level of whiteblood cells at the same level for at least 3 months.
 10. The method ofclaim 1, further comprising administering about 500 mg/day to about 5000mg/day of ACC.
 11. The method of claim 1, wherein the composition isformulated as a pharmaceutical composition or nutraceutical composition,a food supplement, or a medical food.